Therapeutic compounds

ABSTRACT

The invention provides compounds of formula I: 
                         
or a salt thereof as described herein. The invention also provides pharmaceutical compositions comprising a compound of formula I, processes for preparing compounds of formula I, intermediates useful for preparing compounds of formula I and therapeutic methods for treating cancer or treating autoimmune diseases or preventing transplant rejection using compounds of formula I.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.14/373,615, filed Jul. 21, 2014, which is a National Stage Applicationof International Application No. PCT/US2013/022275, filed Jan. 18, 2013,which claims priority to U.S. Provisional Application 61/589,090, filedJan. 20, 2012, the disclosures of which applications are incorporated byreference herein.

BACKGROUND OF THE INVENTION

Monocarboxylate Transporters (MCTs) are members of the solute carrier 16(SLC16) gene family. In mammalian species there are 14 known MCTisoforms and only four (MCT-1, -2, -3, -4) have been demonstrated toperform proton-linked transport of monocarboxylates such as lactate,pyruvate, butyrate, and ketone bodies.

Malignant tumors contain aerobic and hypoxic regions and intratumoralhypoxia increases the risk of cancer advancement, and metastasis. Tumorhypoxia leads to treatment failure, relapse and patient mortality asthese cells are generally resistant to standard chemo- and radiationtherapy. In regions of hypoxia, cancer cells metabolize glucose intolactate, whereas nearby aerobic cancer cells take up this lactate viathe mono-carboxylate transporter 1 (MCT1) for oxidative phosphorylation.MCT1 expression is elevated in an array of human tumors including brain,breast, head, neck, lung and colon.

Under hypoxic conditions, cancer cells upregulate glucose transportersand consume large quantities of glucose. Cancer cells also upregulateglycolytic enzymes and convert glucose into lactate, which is theneffluxed out of the cell via MCT4. The nearby aerobic cancer cells takeup this lactate via the MCT1 for energy generation through oxidativephosphorylation. Thus, the limited glucose available to the tumor isused most efficiently via a synergistic metabolic symbiosis. Thisutilization of lactate as an energy substitute for survival prevents theaerobic cells from consuming large quantities of glucose. Targetedinhibition of MCT4 will lead to lactic acidosis and consequent death ofhypoxic cells, while MCT1 inhibition will cause aerobic cancer cells toconsume glucose instead of lactate, thus resulting in further stress anddeath of hypoxic cancer cells.

The inhibition of MCT1 may also be useful for treating autoimmunediseases and preventing organ transplant rejection. The rate of shortterm graft survival has been improved by current immunosuppressants,however in the past two decades there has been no improvement in theabilities of agents to promote long term graft survival. Recently it hasbeen demonstrated that agents which inhibit MCT1 have been shown to aidprolonged allograft survival, prevent chronic rejection, and inducetolerance in rat allograft models. In an immune response, a rapiddivision of T cells occurs, and these activated T cells use glycolysisas their means for energy production. Interestingly, the activated Tcells use glycolysis even though the cells exist in an aerobicenvironment. As MCT1 plays a critical role in the aerobic glycolysis ofthe activated T cells by exporting lactate, inhibition of MCT1 leads toa buildup of lactate in the cell. This in turn decreases glycolytic fluxeffectively limiting the proliferation of new lymphocytes.

There is a current need for agents that are useful for treating orpreventing cancer or that are useful for treating or preventingautoimmune diseases or that are useful for preventing transplantrejection. There is also a need for agents that are useful fordiagnosing cancer or autoimmune diseases or for imaging cancerous cellsor cells involved in an autoimmune response.

SUMMARY OF THE INVENTION

In one embodiment, the invention provides a compound of the inventionwhich is a compound of formula I:

wherein:

X is CN, Y is R⁵ and the dashed bond is absent; or

X is —C(═O)—, Y is O and the dashed bond is a single bond;

R¹ is H or (C₁-C₆)alkyl;

R² is H, (C₁-C₆)alkyl, (C₁-C₆)alkoxy, halo(C₁-C₆)alkyl, halo, —OH, —CN,—NO₂, —CO₂R_(a), —C(═O)R_(a), —NR_(a)(C═O)R_(b), —C(═O)NR_(c)R_(d) or—NR_(c)R_(d);

R³ is H, (C₁-C₆)alkyl, (C₁-C₆)alkoxy, halo(C₁-C₆)alkyl, halo, —OH, —CN,—NO₂, —CO₂R_(a), —C(═O)R_(a), —NR_(a)(C═O)R_(b), —C(═O)NR_(c)R_(d) or—NR_(c)R_(d);

R⁴ is H, (C₁-C₆)alkyl, (C₁-C₆)alkoxy, halo(C₁-C₆)alkyl, halo, —OH, —CN,—NO₂, —CO₂R_(a), —C(═O)R_(a), —NR_(a)(C═O)R_(b), —C(═O)NR_(c)R_(d) or—NR_(c)R_(d);

R⁵ is H, (C₁-C₆)alkyl, (C₁-C₆)alkoxy, halo(C₁-C₆)alkyl, halo, —OH, —CN,—NO₂, —CO₂R_(a), —C(═O)R_(a), —NR_(a)(C═O)R_(b), —C(═O)NR_(c)R_(d) or—NR_(c)R_(d);

R^(6a) and R^(6b) are each independently H, (C₁-C₇)alkyl,(C₂-C₇)alkenyl, (C₂-C₇)alkynyl, aryl(C₁-C₆)alkyl-,heteroaryl(C₁-C₆)alkyl-, heterocycle(C₁-C₆)alkyl-, aryl, heterocycle orheteroaryl, wherein any (C₁-C₇)alkyl, (C₂-C₇)alkenyl, (C₂-C₇)alkynyl,aryl(C₁-C₆)alkyl-, heteroaryl(C₁-C₆)alkyl-, heterocycle(C₁-C₆)alkyl-,aryl, heterocycle or heteroaryl of R^(6a) and R^(6b) is optionallysubstituted with one or more (e.g. 1, 2, 3, 4 or 5) Z¹ groups; or R^(6a)and R^(6b) together with the nitrogen to which they are attached form aheterocycle, wherein the heterocycle is optionally substituted with oneor more (e.g. 1, 2, 3, 4 or 5) Z¹ groups;

each R_(a) is independently H, (C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl;

each R_(b) is independently (C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl;

R_(c) and R_(d) are each independently H, (C₁-C₆)alkyl,aryl(C₁-C₆)alkyl- or aryl; or R_(c) and R_(d) together with the nitrogento which they are attached form a pyrrolidino, piperidino, piperazino,azetidino, morpholino, or thiomorpholino;

each Z¹ is independently (C₁-C₆)alkyl, (C₁-C₆)alkoxy, halo(C₁-C₆)alkyl,halo, —OH, —CN, —NO₂, —CO₂R_(e), —C(═O)R_(e), —NR_(e)(C═O)R_(f),—C(═O)NR_(g)R_(h) or —NR_(g)R_(h);

each R_(e) is independently H, (C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl;

each R_(f) is independently (C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl; and

R_(g) and R_(h) are each independently H, (C₁-C₆)alkyl,aryl(C₁-C₆)alkyl- or aryl; or R_(g) and R_(h) together with the nitrogento which they are attached form a pyrrolidino, piperidino, piperazino,azetidino, morpholino, or thiomorpholino;

or a salt thereof.

The invention also provides a pharmaceutical composition comprising acompound of formula I, or a pharmaceutically acceptable salt thereof,and a pharmaceutically acceptable diluent or carrier.

The invention also provides a method for inhibiting cancer (e.g. lung,breast, brain, prostate, pancreatic, colorectal, ovarian, head and neck)cell growth comprising contacting the cancer cell in vitro or in vivowith an effective amount of a compound of formula I, or a salt thereof.

The invention also provides a method for treating cancer (e.g. lung,breast, brain, prostate, pancreatic, colorectal, ovarian, head and neckcancer) in a mammal (e.g. a human) comprising administering to themammal an effective amount of compound as described in formula I, or apharmaceutically acceptable salt thereof.

The invention also provides a method for treating an autoimmune disease(e.g. rheumatoid arthritis) in a mammal (e.g. a human) comprisingadministering to the mammal an effective amount of a compound of formulaI, or a pharmaceutically acceptable salt thereof.

The invention also provides a method for preventing transplant rejection(e.g. heart, kidney, eye, liver, lung, pancreas, intestine, and thymustransplant rejection) or tissue graft rejection (e.g. bone, tendon,cornea, skin, heart valve and vein) in a mammal (e.g. a human)comprising administering to the mammal an effective amount of a compoundof formula I, or a pharmaceutically acceptable salt thereof.

The invention also provides a method for diagnosing cancer (e.g. lung,breast, brain, prostate, pancreatic, colorectal, ovarian, head and neckcancer) or for diagnosing an autoimmune disease (e.g. rheumatoidarthritis) in a mammal comprising administering to the mammal (e.g. ahuman) an effective amount of a compound of formula I, or apharmaceutically acceptable salt thereof and measuring or imaging thefluorescence of the compound of formula I wherein the fluorescencecorrelates with cancer or an autoimmune disease.

The invention also provides a method for imaging cancerous cells (e.g.brain, breast, head, neck, lung or colon cancer cells) or for imagingcells involved in an autoimmune response (e.g. rheumatoid arthritis) orfor imaging cells involved in transplant rejection (e.g. heart, kidney,eye, liver, lung, pancreas, intestine, and thymus transplant rejection)or tissue graft rejection (e.g. bone, tendon, cornea, skin, heart valveand vein) comprising contacting the cells in vivo or in vitro with acompound of formula I, or a salt thereof and imaging the fluorescence ofthe compound of formula I while in contact with the cell.

The invention also provides a method for altering brain function (e.g.long-term memory formation) in a mammal (e.g. a human) comprisingadministering to the mammal an effective amount of a compound of formulaI, or a pharmaceutically acceptable salt thereof.

The invention also provides a compound of formula I, or apharmaceutically acceptable salt thereof, for use in medical therapy.

The invention also provides the use of a compound of formula I, or apharmaceutically acceptable salt thereof, for the manufacture of amedicament useful for the treatment of cancer (e.g. lung, breast, brain,prostate, pancreatic, colorectal, ovarian, head and neck cancer) in amammal (e.g. a human).

The invention also provides a compound of formula I, or apharmaceutically acceptable salt thereof, for use in the prophylactic ortherapeutic treatment of cancer (e.g. lung, breast, brain, prostate,pancreatic, colorectal, ovarian, head and neck cancer).

The invention also provides a compound of formula I, or apharmaceutically acceptable salt thereof, for use in the prophylactic ortherapeutic alteration of brain function

The invention also provides the use of a compound of formula I, or apharmaceutically acceptable salt thereof, for the manufacture of amedicament useful for the treatment of an autoimmune disease (e.g.rheumatoid arthritis) in a mammal (e.g. a human).

The invention also provides the use of a compound of formula I, or apharmaceutically acceptable salt thereof, for the manufacture of amedicament useful for altering brain function (e.g. long-term memoryformation) in a mammal (e.g. a human).

The invention also provides a compound of formula I, or apharmaceutically acceptable salt thereof, for use in the prophylactic ortherapeutic treatment of an autoimmune disease (e.g. rheumatoidarthritis).

The invention also provides the use of a compound of formula I, or apharmaceutically acceptable salt thereof, for the manufacture of amedicament useful for preventing transplant rejection (e.g. heart,kidney, eye, liver, lung, pancreas, intestine, and thymus transplantrejection) or tissue graft rejection (e.g. bone, tendon, cornea, skin,heart valve and vein) in a mammal (e.g. a human).

The invention also provides a compound of formula I, or apharmaceutically acceptable salt thereof, for use in the prevention oftransplant rejection (e.g. heart, kidney, eye, liver, lung, pancreas,intestine, and thymus transplant rejection) or tissue graft rejection(e.g. bone, tendon, cornea, skin, heart valve and vein).

The invention also provides a compound of formula I, or a salt thereoffor use in diagnosing cancer (e.g. lung, breast, brain, prostate,pancreatic, colorectal, ovarian, head and neck) or for diagnosing aautoimmune disease (e.g. rheumatoid arthritis).

The invention also provides a compound of formula I, or a salt thereoffor use in imaging cancerous cells (e.g. brain, breast, head, neck, lungor colon cancer cells) or for imaging cells involved in an autoimmuneresponse (e.g. rheumatoid arthritis) or for imaging cells involved intransplant rejection (e.g. heart, kidney, eye, liver, lung, pancreas,intestine, and thymus transplant rejection) or tissue graft rejection(e.g. bone, tendon, cornea, skin, heart valve and vein).

The invention also provides processes and intermediates disclosed hereinthat are useful for preparing compounds of formula I or salts thereof.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates the anti-cancer effect of Compound 20, and Compound14 on the tumor growth in WiDr xenograft tumor model from Example 7.

FIG. 2 illustrates the anti-cancer effect of Compound 20, and Compound14 on the tumor growth in WiDr xenograft tumor model from Example 7.

FIG. 3 illustrates the Anti-Cancer Effect of Compound 20 on the TumorGrowth in the WiDr xenograft tumor model (after 14 Days) from Example 8.

FIG. 4 illustrates the anti-cancer effect of Compound 20 on the tumorgrowth in the WiDr xenograft tumor model (after 22 Days) from Example 8.

FIG. 5 illustrates the anti-cancer effect of Compound 20 on the tumorweight in the WiDr xenograft tumor model from Example 8.

FIG. 6 illustrates the body weight changes in mice from Example 8.

DETAILED DESCRIPTION

The following definitions are used, unless otherwise described.

The term “halo” or “halogen” as used herein refers to fluoro, chloro,bromo, or iodo.

The term “alkyl” as used herein refers to straight and branchedhydrocarbon groups; the term “alkenyl” as used herein refers to straightor branched hydrocarbon groups containing at least one carbon-carbondouble bond; the term “alkynyl” as used herein refers to straight orbranched hydrocarbon groups containing at least one carbon-carbon triplebond; the term “alkoxy” as used herein refers to groups of the formulaalkyl-O—, where alkyl is as defined herein. Reference to an individualradical such as propyl embraces only the straight chain radical, abranched chain isomer such as isopropyl being specifically referred to.

The term “haloalkyl” as used herein refers to an alkyl as definedherein, wherein one or more hydrogen atoms are each replaced by a halosubstituent. For example, a halo (C₁-C₆)alkyl is a (C₁-C₆)alkyl whereinone or more of the hydrogen atoms have been replaced by a halosubstituent. Such a range includes one halo substituent on the alkylgroup to complete halogenation of the alkyl group.

The term “aryl” as used herein refers to a single aromatic ring or amultiple condensed ring system. For example, an aryl group can have 6 to20 carbon atoms, 6 to 14 carbon atoms, or 6 to 12 carbon atoms. Arylincludes a phenyl radical. Aryl also includes multiple condensed ringsystems (e.g. ring systems comprising 2, 3 or 4 rings) having about 9 to20 carbon atoms in which at least one ring is aromatic. Such multiplecondensed ring systems may be optionally substituted with one or more(e.g. 1, 2 or 3) oxo groups on any carbocycle portion of the multiplecondensed ring system. It is to be understood that the point ofattachment of a multiple condensed ring system, as defined above, can beat any position of the ring system including an aryl or a carbocycleportion of the ring. Typical aryl groups include, but are not limitedto, phenyl, indenyl, naphthyl, 1,2,3,4-tetrahydronaphthyl, anthracenyl,and the like.

The term “heteroaryl” as used herein refers to a single aromatic ring ora multiple condensed ring system. The term includes single aromaticrings of from about 1 to 6 carbon atoms and about 1-4 heteroatomsselected from the group consisting of oxygen, nitrogen and sulfur in therings. The sulfur and nitrogen atoms may also be present in an oxidizedform provided the ring is aromatic. Such rings include but are notlimited to pyridyl, pyrimidinyl, oxazolyl or furyl. The term alsoincludes multiple condensed ring systems (e.g. ring systems comprising2, 3 or 4 rings) wherein a heteroaryl group, as defined above, can becondensed with one or more heteroaryls (e.g. naphthyridinyl),heterocycles, (e.g. 1,2,3,4-tetrahydronaphthyridinyl), carbocycles (e.g.5,6,7,8-tetrahydroquinolyl) or aryls (e.g. indazolyl) to form a multiplecondensed ring system. Such multiple condensed ring systems may beoptionally substituted with one or more (e.g. 1, 2, 3 or 4) oxo groupson the carbocycle or heterocycle portions of the condensed ring. It isto be understood that the point of attachment of a multiple condensedring system (as defined above for a heteroaryl) can be at any positionof the multiple condensed ring system including a heteroaryl,heterocycle, aryl or carbocycle portion of the multiple condensed ringsystem and at any suitable atom of the multiple condensed ring systemincluding a carbon atom and heteroatom (e.g. a nitrogen). Exemplaryheteroaryls include but are not limited to pyridyl, pyrrolyl, pyrazinyl,pyrimidinyl, pyridazinyl, pyrazolyl, thienyl, indolyl, imidazolyl,oxazolyl, thiazolyl, furyl, oxadiazolyl, thiadiazolyl, quinolyl,isoquinolyl, benzothiazolyl, benzoxazolyl, indazolyl, quinoxalyl,quinazolyl, 5,6,7,8-tetrahydroisoquinolinyl benzofuranyl, benzimidazolyland thianaphthenyl.

The term “heterocyclyl” or “heterocycle” as used herein refers to asingle saturated or partially unsaturated ring or a multiple condensedring system. The term includes single saturated or partially unsaturatedrings (e.g. 3, 4, 5, 6 or 7-membered rings) from about 1 to 6 carbonatoms and from about 1 to 3 heteroatoms selected from the groupconsisting of oxygen, nitrogen and sulfur in the ring. The ring may besubstituted with one or more (e.g. 1, 2 or 3) oxo groups and the sulfurand nitrogen atoms may also be present in their oxidized forms. Suchrings include but are not limited to azetidinyl, tetrahydrofuranyl orpiperidinyl. The term “heterocycle” also includes multiple condensedring systems (e.g. ring systems comprising 2, 3 or 4 rings) wherein asingle heterocycle ring (as defined above) can be condensed with one ormore heterocycles (e.g. decahydronapthyridinyl), carbocycles (e.g.decahydroquinolyl) or aryls. The rings of a multiple condensed ringsystem can be connected to each other via fused, spiro and bridged bondswhen allowed by valency requirements. It is to be understood that thepoint of attachment of a multiple condensed ring system (as definedabove for a heterocycle) can be at any position of the multiplecondensed ring system including a heterocycle, aryl and carbocycleportion of the ring. It is also to be understood that the point ofattachment for a heterocycle or heterocycle multiple condensed ringsystem can be at any suitable atom of the heterocycle or heterocyclemultiple condensed ring system including a carbon atom and a heteroatom(e.g. a nitrogen). Exemplary heterocycles include, but are not limitedto aziridinyl, azetidinyl, pyrrolidinyl, piperidinyl, homopiperidinyl,morpholinyl, thiomorpholinyl, piperazinyl, tetrahydrofuranyl,dihydrooxazolyl, tetrahydropyranyl, tetrahydrothiopyranyl,1,2,3,4-tetrahydroquinolyl, benzoxazinyl, dihydrooxazolyl, chromanyl,1,2-dihydropyridinyl, 2,3-dihydrobenzofuranyl, 1,3-benzodioxolyl and1,4-benzodioxanyl.

The term “carbocycle” or “carbocyclyl” refers to a single saturated(i.e., cycloalkyl) or a single partially unsaturated (e.g.,cycloalkenyl, cycloalkadienyl, etc.) ring having 3 to 8 carbon atoms(i.e. (C₃-C₈)carbocycle). The term “carbocycle” or “carbocyclyl” alsoincludes multiple condensed ring systems (e.g. ring systems comprising2, 3 or 4 carbocyclic rings). Accordingly, carbocycle includesmulticyclic carbocyles having 7 to 12 carbon atoms as a bicycle, and upto about 20 carbon atoms as a polycycle. Multicyclic carbocyles can beconnected to each other via a single carbon atom to form a spiroconnection (e.g. spiropentane, spiro[4,5]decane, spiro[4.5]decane, etc),via two adjacent carbon atoms to form a fused connection such as abicyclo [4,5], [5,5], [5,6] or [6,6] system, or 9 or 10 ring atomsarranged as a bicyclo [5,6] or [6,6] system (e.g. decahydronaphthalene,norsabinane, norcarane) or via two non-adjacent carbon atoms to form abridged connection (e.g. norbornane, bicyclo[2.2.2]octane, etc). The“carbocycle” or “carbocyclyl” can also be optionally substituted withone or more (e.g. 1, 2 or 3) oxo groups. Non-limiting examples ofmonocyclic carbocycles include cyclopropyl, cyclobutyl, cyclopentyl,1-cyclopent-1-enyl, 1-cyclopent-2-enyl, 1-cyclopent-3-enyl, cyclohexyl,1-cyclohex-1-enyl, 1-cyclohex-2-enyl and 1-cyclohex-3-enyl.

When trade names are used herein, applicants intend to independentlyinclude the tradename product and the active pharmaceuticalingredient(s) of the tradename product.

It will be appreciated by those skilled in the art that compounds of theinvention having a chiral center may exist in and be isolated inoptically active and racemic forms. Some compounds may exhibitpolymorphism. It is to be understood that the present inventionencompasses any racemic, optically-active, polymorphic, orstereoisomeric form, or mixtures thereof, of a compound of theinvention, which possess the useful properties described herein, itbeing well known in the art how to prepare optically active forms (forexample, by resolution of the racemic form by recrystallizationtechniques, by synthesis from optically-active starting materials, bychiral synthesis, or by chromatographic separation using a chiralstationary phase.

Specific values listed below for radicals, substituents, and ranges, arefor illustration only; they do not exclude other defined values or othervalues within defined ranges for the radicals and substituents.

A specific group of compounds of formula I are compounds of formula Ia:

or a salt thereof.

A specific group of compounds of formula I are compounds of formula Ib:

or a salt thereof.

Another specific group of compounds of formula I are compounds offormula Ic:

or a salt thereof.

Specific values listed below are values for compounds of formula I aswell as all sub-formulas of formula I (e.g. formulas Ia, Ib, Ic, 1b, 2band 3b).

Specifically, (C₁-C₆)alkyl can be methyl, ethyl, propyl, isopropyl,butyl, iso-butyl, sec-butyl, pentyl, 3-pentyl or hexyl; (C₁-C₆)alkoxycan be methoxy, ethoxy, propoxy, isopropoxy, butoxy, iso-butoxy,sec-butoxy, pentoxy, 3-pentoxy, or hexyloxy; (C₂-C₆)alkenyl can bevinyl, allyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 3-butenyl,1,-pentenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl, 1-hexenyl, 2-hexenyl,3-hexenyl, 4-hexenyl, or 5-hexenyl; (C₂-C₆)alkynyl can be ethynyl,1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1-pentynyl,2-pentynyl, 3-pentynyl, 4-pentynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl,4-hexynyl, or 5-hexynyl; and halo(C₁-C₆)alkyl can be iodomethyl,bromomethyl, chloromethyl, fluoromethyl, trifluoromethyl, 2-chloroethyl,2-fluoroethyl, 2,2,2-trifluoroethyl, or pentafluoroethyl.

A specific value for R¹ is H.

A specific value for R² is H or methoxy.

Another specific value for R² is methoxy.

Another specific value for R² is H.

A specific value for R³ is H.

A specific value for each of R^(6a) and R^(6b) independently is(C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl, aryl(C₁-C₆)alkyl-, aryl orheterocycle, wherein any (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl,aryl(C₁-C₆)alkyl-, aryl or heterocycle of R^(6a) and R^(6b) isoptionally substituted with one or more Z¹ groups; or R^(6a) and R^(6b)together with the nitrogen to which they are attached form a heterocycleoptionally substituted with one or more Z¹ groups.

Another specific independent value for each of R^(6a) and R^(6b)independently is (C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl, wherein any(C₁-C₆)alkyl, aryl(C₁-C₆)alkyl- or aryl of R^(6a) and R^(6b) isoptionally substituted with one or more Z¹ groups.

A specific value for —NR^(6a)R^(6b) is:

Another specific value for —NR^(6a)R^(6b) is:

Another specific value for —NR^(6a)R^(6b) is:

Another specific value for —NR^(6a)R^(6b) is

A specific compound of formula I is:

or a salt thereof.

A specific compound of formula I is:

or a salt thereof.

Another specific compound of formula I is:

or a salt thereof.

Another specific compound of formula I is:

or a salt thereof.

Another specific compound of formula I is:

or a salt thereof.

Another specific compound of formula I is:

or a salt thereof.

In one embodiment of the invention the compounds of formula I do notinclude:

-   (E)-2-cyano-3-(4-(dibutylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(dipropylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(dimethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-2-methylphenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-2-methoxyphenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-5-methoxy-2-methylphenyl)acrylic    acid,-   (E)-2-cyano-3-(4-(di-p-tolylamino)phenyl)acrylic acid-   (E)-3-(2-acetamido-4-(diethylamino)-5-methoxyphenyl)-2-cyanoacrylic    acid,-   (E)-2-cyano-3-(4-(dimethylamino)-2-methylphenyl)acrylic acid,-   (E)-3-(2-chloro-4-(dimethylamino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-aminophenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(chloromethyl)amino)phenyl)-2-cyanoacrylic    acid,-   (E)-3-(4-(bis(2-methoxyethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-(bis(2-chloroethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(methyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-2-cyano-3-(4-(diphenylamino)phenyl)acrylic acid,-   (E)-3-(4-(bis(4-(dimethylamino)phenyl)amino)phenyl)-2-cyanoacrylic    acid,-   7-(dimethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(diethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(dibutylamino)-2-oxo-2H-chromene-3-carboxylic acid or-   7-(diphenylamino)-2-oxo-2H-chromene-3-carboxylic acid.

In another embodiment of the invention the compounds of formula I do notinclude:

-   (E)-2-cyano-3-(4-(dipropylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(dimethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-2-methylphenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-2-methoxyphenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-5-methoxy-2-methylphenyl)acrylic    acid,-   (E)-2-cyano-3-(4-(di-p-tolylamino)phenyl)acrylic acid,-   (E)-3-(2-acetamido-4-(diethylamino)-5-methoxyphenyl)-2-cyanoacrylic    acid,-   (E)-2-cyano-3-(4-(dimethylamino)-2-methylphenyl)acrylic acid,-   (E)-3-(2-chloro-4-(dimethylamino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-aminophenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(chloromethyl)amino)phenyl)-2-cyanoacrylic    acid,-   (E)-3-(4-(bis(2-methoxyethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-(bis(2-chloroethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(methyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-2-cyano-3-(4-(diphenylamino)phenyl)acrylic acid or-   (E)-3-(4-(bis(4-(dimethylamino)phenyl)amino)phenyl)-2-cyanoacrylic    acid.

In another embodiment of the invention the compounds of formula I do notinclude:

-   7-(dimethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(diethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(dibutylamino)-2-oxo-2H-chromene-3-carboxylic acid or    7-(diphenylamino)-2-oxo-2H-chromene-3-carboxylic acid.

In another embodiment of the invention the compounds of formula I do notinclude:

-   (E)-2-cyano-3-(4-(dipropylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(dimethylamino)phenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-2-methylphenyl)acrylic acid,-   (E)-2-cyano-3-(4-(diethylamino)-5-methoxy-2-methylphenyl)acrylic    acid,-   (E)-2-cyano-3-(4-(di-p-tolylamino)phenyl)acrylic acid,-   (E)-3-(2-acetamido-4-(diethylamino)-5-methoxyphenyl)-2-cyanoacrylic    acid,-   (E)-2-cyano-3-(4-(dimethylamino)-2-methylphenyl)acrylic acid,-   (E)-3-(2-chloro-4-(dimethylamino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-aminophenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(chloromethyl)amino)phenyl)-2-cyanoacrylic    acid,-   (E)-3-(4-(bis(2-methoxyethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-(bis(2-chloroethyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-3-(4-((2-chloroethyl)(methyl)amino)phenyl)-2-cyanoacrylic acid,-   (E)-2-cyano-3-(4-(diphenylamino)phenyl)acrylic acid,-   (E)-3-(4-(bis(4-(dimethylamino)phenyl)amino)phenyl)-2-cyanoacrylic    acid,-   7-(dimethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(diethylamino)-2-oxo-2H-chromene-3-carboxylic acid,-   7-(dibutylamino)-2-oxo-2H-chromene-3-carboxylic acid or-   7-(diphenylamino)-2-oxo-2H-chromene-3-carboxylic acid.

In another embodiment of the invention the compounds of formula I do notinclude:

-   (E)-2-cyano-3-(4-(diethylamino)-2-methoxyphenyl)acrylic acid.

Processes for preparing compounds of formula I are provided as furtherembodiments of the invention and are illustrated in Schemes 1-3 in whichthe meanings of the generic radicals are as given above unless otherwisequalified. Scheme 1 depicts a general procedure for the synthesis of2-cyano-phenyl acrylic acid compounds of formula I, Scheme 2 depicts ageneral procedure for the synthesis of methoxy substituted2-cyano-phenyl acrylic acid compounds of formula I and Scheme 3 depictsa general procedure for the synthesis of 2-oxo-2H-chromene-3-carboxylicacid compounds of formula I. An intermediate useful for preparing acompound of formula I, is a compound of the formula 1a, 2a or 3a or asalt thereof.

In cases where compounds are sufficiently basic or acidic, a salt of acompound of formula I can be useful as an intermediate for isolating orpurifying a compound of formula I. Additionally, administration of acompound of formula I as a pharmaceutically acceptable acid or base saltmay be appropriate. Examples of pharmaceutically acceptable salts areorganic acid addition salts formed with acids which form a physiologicalacceptable anion, for example, tosylate, methanesulfonate, acetate,citrate, malonate, tartarate, succinate, benzoate, ascorbate,α-ketoglutarate, and α-glycerophosphate. Suitable inorganic salts mayalso be formed, including hydrochloride, sulfate, nitrate, bicarbonate,and carbonate salts.

Pharmaceutically acceptable salts may be obtained using standardprocedures well known in the art, for example by reacting a sufficientlybasic compound such as an amine with a suitable acid affording aphysiologically acceptable anion. Alkali metal (for example, sodium,potassium or lithium) or alkaline earth metal (for example calcium)salts of carboxylic acids can also be made.

The compounds of formula I can be formulated as pharmaceuticalcompositions and administered to a mammalian host, such as a humanpatient in a variety of forms adapted to the chosen route ofadministration, i.e., orally or parenterally, by intravenous,intramuscular, topical or subcutaneous routes.

Thus, the present compounds may be systemically administered, e.g.,orally, in combination with a pharmaceutically acceptable vehicle suchas an inert diluent or an assimilable edible carrier. They may beenclosed in hard or soft shell gelatin capsules, may be compressed intotablets, or may be incorporated directly with the food of the patient'sdiet. For oral therapeutic administration, the active compound may becombined with one or more excipients and used in the form of ingestibletablets, buccal tablets, troches, capsules, elixirs, suspensions,syrups, wafers, and the like. Such compositions and preparations shouldcontain at least 0.1% of active compound. The percentage of thecompositions and preparations may, of course, be varied and mayconveniently be between about 2 to about 60% of the weight of a givenunit dosage form. The amount of active compound in such therapeuticallyuseful compositions is such that an effective dosage level will beobtained.

The tablets, troches, pills, capsules, and the like may also contain thefollowing: binders such as gum tragacanth, acacia, corn starch orgelatin; excipients such as dicalcium phosphate; a disintegrating agentsuch as corn starch, potato starch, alginic acid and the like; alubricant such as magnesium stearate; and a sweetening agent such assucrose, fructose, lactose or aspartame or a flavoring agent such aspeppermint, oil of wintergreen, or cherry flavoring may be added. Whenthe unit dosage form is a capsule, it may contain, in addition tomaterials of the above type, a liquid carrier, such as a vegetable oilor a polyethylene glycol. Various other materials may be present ascoatings or to otherwise modify the physical form of the solid unitdosage form. For instance, tablets, pills, or capsules may be coatedwith gelatin, wax, shellac or sugar and the like. A syrup or elixir maycontain the active compound, sucrose or fructose as a sweetening agent,methyl and propylparabens as preservatives, a dye and flavoring such ascherry or orange flavor. Of course, any material used in preparing anyunit dosage form should be pharmaceutically acceptable and substantiallynon-toxic in the amounts employed. In addition, the active compound maybe incorporated into sustained-release preparations and devices.

The active compound may also be administered intravenously orintraperitoneally by infusion or injection. Solutions of the activecompound or its salts can be prepared in water, optionally mixed with anontoxic surfactant. Dispersions can also be prepared in glycerol,liquid polyethylene glycols, triacetin, and mixtures thereof and inoils. Under ordinary conditions of storage and use, these preparationscontain a preservative to prevent the growth of microorganisms.

The pharmaceutical dosage forms suitable for injection or infusion caninclude sterile aqueous solutions or dispersions or sterile powderscomprising the active ingredient which are adapted for theextemporaneous preparation of sterile injectable or infusible solutionsor dispersions, optionally encapsulated in liposomes. In all cases, theultimate dosage form should be sterile, fluid and stable under theconditions of manufacture and storage. The liquid carrier or vehicle canbe a solvent or liquid dispersion medium comprising, for example, water,ethanol, a polyol (for example, glycerol, propylene glycol, liquidpolyethylene glycols, and the like), vegetable oils, nontoxic glycerylesters, and suitable mixtures thereof. The proper fluidity can bemaintained, for example, by the formation of liposomes, by themaintenance of the required particle size in the case of dispersions orby the use of surfactants. The prevention of the action ofmicroorganisms can be brought about by various antibacterial andantifungal agents, for example, parabens, chlorobutanol, phenol, sorbicacid, thimerosal, and the like. In many cases, it will be preferable toinclude isotonic agents, for example, sugars, buffers or sodiumchloride. Prolonged absorption of the injectable compositions can bebrought about by the use in the compositions of agents delayingabsorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the activecompound in the required amount in the appropriate solvent with variousof the other ingredients enumerated above, as required, followed byfilter sterilization. In the case of sterile powders for the preparationof sterile injectable solutions, the preferred methods of preparationare vacuum drying and the freeze drying techniques, which yield a powderof the active ingredient plus any additional desired ingredient presentin the previously sterile-filtered solutions.

For topical administration, the present compounds may be applied in pureform, i.e., when they are liquids. However, it will generally bedesirable to administer them to the skin as compositions orformulations, in combination with a dermatologically acceptable carrier,which may be a solid or a liquid.

Useful solid carriers include finely divided solids such as talc, clay,microcrystalline cellulose, silica, alumina and the like. Useful liquidcarriers include water, alcohols or glycols or water-alcohol/glycolblends, in which the present compounds can be dissolved or dispersed ateffective levels, optionally with the aid of non-toxic surfactants.Adjuvants such as fragrances and additional antimicrobial agents can beadded to optimize the properties for a given use. The resultant liquidcompositions can be applied from absorbent pads, used to impregnatebandages and other dressings, or sprayed onto the affected area usingpump-type or aerosol sprayers.

Thickeners such as synthetic polymers, fatty acids, fatty acid salts andesters, fatty alcohols, modified celluloses or modified mineralmaterials can also be employed with liquid carriers to form spreadablepastes, gels, ointments, soaps, and the like, for application directlyto the skin of the user.

Examples of useful dermatological compositions which can be used todeliver the compounds of formula I to the skin are known to the art; forexample, see Jacquet et al. (U.S. Pat. No. 4,608,392), Geria (U.S. Pat.No. 4,992,478), Smith et al. (U.S. Pat. No. 4,559,157) and Wortzman(U.S. Pat. No. 4,820,508).

Useful dosages of the compounds of formula I can be determined bycomparing their in vitro activity, and in vivo activity in animalmodels. Methods for the extrapolation of effective dosages in mice, andother animals, to humans are known to the art; for example, see U.S.Pat. No. 4,938,949.

The amount of the compound, or an active salt or derivative thereof,required for use in treatment will vary not only with the particularsalt selected but also with the route of administration, the nature ofthe condition being treated and the age and condition of the patient andwill be ultimately at the discretion of the attendant physician orclinician.

In general, however, a suitable dose will be in the range of from about0.5 to about 100 mg/kg, e.g., from about 10 to about 75 mg/kg of bodyweight per day, such as 3 to about 50 mg per kilogram body weight of therecipient per day, preferably in the range of 6 to 90 mg/kg/day, mostpreferably in the range of 15 to 60 mg/kg/day.

The compound is conveniently formulated in unit dosage form; forexample, containing 5 to 1000 mg, conveniently 10 to 750 mg, mostconveniently, 50 to 500 mg of active ingredient per unit dosage form. Inone embodiment, the invention provides a composition comprising acompound of the invention formulated in such a unit dosage form.

The desired dose may conveniently be presented in a single dose or asdivided doses administered at appropriate intervals, for example, astwo, three, four or more sub-doses per day. The sub-dose itself may befurther divided, e.g., into a number of discrete loosely spacedadministrations; such as multiple inhalations from an insufflator or byapplication of a plurality of drops into the eye.

Aggressive glycolysis is the hallmark of all advanced stage tumors.Accordingly, compounds and compositions described herein may be usefulto treat cancers that express MCT's (e.g. lung, breast, brain, prostate,pancreatic, colorectal, ovarian, head and neck).

In addition, compounds and compositions described herein may be usefulto prevent organ (e.g. heart, kidney, eye, liver, lung, pancreas,intestine, and thymus) transplant rejection as well as tissues graft(e.g. bone, tendon, cornea, skin, heart valves, and veins) rejection.Compounds and compositions described herein may also be useful fortreating autoimmune diseases (e.g. rheumatoid arthritis).

Compounds of the invention can also be administered in combination withother therapeutic agents, for example, other agents that are useful forthe treatment of cancer (e.g. lung, breast, brain, prostate, pancreatic,colorectal, ovarian, head and neck cancer) or autoimmune diseases (e.g.rheumatoid arthritis) or agents that are useful for preventingtransplant rejection such as organ transplant (e.g. heart, kidney, eye,liver, lung, pancreas, intestine, and thymus) rejection and or tissuegraft (e.g. bone, tendon, cornea, skin, heart valve, and vein)rejection. Accordingly, in one embodiment the invention also provides acomposition comprising a compound of formula I, or a pharmaceuticallyacceptable salt thereof, at least one other therapeutic agent, and apharmaceutically acceptable diluent or carrier. The invention alsoprovides a kit comprising a compound of formula I, or a pharmaceuticallyacceptable salt thereof, at least one other therapeutic agent, packagingmaterial, and instructions for administering the compound of formula Ior the pharmaceutically acceptable salt thereof and the othertherapeutic agent or agents to an animal to treat cancer or anautoimmune disease or to prevent transplant rejection.

The ability of a compound of the invention to inhibit MCT1 may bedetermined using pharmacological models which are well known to the art,or using Test A described below.

Test A. MCT1 Inhibition Via Lactate Uptake Rat Brain Endothelial Cells

The entire lactate uptake studies for the inhibition of MCT1 werecarried out on RBE4 (Rat Brain Endothelial 4) cells. The expression ofMCT1 on these cells was confirmed by Western Blotting. The cells wereplated approximately 20-24 hours before the experiment, the number ofcells being approximately 10⁵ cells per well. The test compounds weredissolved in DMSO and diluted 1000 times using a solution of HEPESbuffer (pH 7.43) which consists of 3 μM ¹⁴C-Lactate and 2 μM L-Lactate.The cells were washed twice with 500 μL of HEPES buffer and the cellswere allowed to equilibrate for 15 minutes at 37° C. The HEPES bufferwas removed and 250 μL of the test sample was added in triplicates. Thiswas repeated for all the compounds, including the controls (CHC andDMSO). After 15 minutes, the compounds were removed from the well and500 μL of ice-cold stop buffer (0.1 mM CHC solution in HEPES buffer) wasadded. The plate was kept on ice. Now, the HEPES buffer in onetriplicate was removed and DMSO solution was added and immediatelyremoved and ice-cold stop buffer was added. This was considered as“Zero”. One triplicate was left blank, which was used for protein assayafter lysing the cells. The cells were washed twice with ice-cold HEPESbuffer and then 250 μL of 0.1M NaOH in 5% Triton-X solution was addedand the plate is kept on a shaker for 40 minutes to lyse the cells. 150μL of the lysed cells was added into 4 mL of the scintillation fluid ina scintillation vial and scintillation count was obtained indisintegrations per minute (dpm). The percent inhibition values werecalculated by taking DMSO as minimum. Concentration study (usually 9-12dilutions) was done to determine the IC₅₀ of each compound.

Experimental results from Test A including results for representativecompounds of the invention are shown in Table 1. These resultsdemonstrate that compounds of the invention inhibit MCT1. Accordinglycompounds of the invention may be useful as therapeutic agents for thetreatment of cancer (lung, breast, brain, prostate, pancreatic,colorectal, ovarian, head or neck cancer) or autoimmune diseases (e.g.rheumatoid arthritis) or for preventing transplant rejection (e.g.heart, kidney, eye, liver, lung, pancreas, intestine, or thymustransplant rejection) or for preventing tissue graft rejection (e.g.bone, tendon, cornea, skin, heart valve or vein graft rejection).

Compounds of the inventions may also be useful for diagnosing cancer orcells involved in autoimmune diseases or transplant rejection or tissuegraft rejection. The coumarin compounds described herein exhibitfluorescence of excitation ˜350 nm and emission ˜450 nm. Since rapidlygrowing cells such as cancer cells and T cells uptake these compoundsmore than normal cells, the fluorescent MCT1 inhibitors described hereinmay be useful for imaging cancerous cells or cells involved in anautoimmune response. Additionally, compounds of the invention may beuseful as pharmacological tools for the further investigation of cancerfunction or for the processes involved in an autoimmune response.

TABLE 1 Compound IC₅₀ (μM) Number Compound (average)  4

0.026  5

0.12  6

0.95  7

0.088  8

0.34  9

0.066 10

0.043 11

0.064 12

0.11 13

0.0095 14

0.0076 15

0.023 16

0.025 17

0.091 18

0.015 19

0.019 20

0.008 21

0.059 22

0.299 23

0.25 24

0.15 25

0.048 26

0.22 27

0.20 28

0.92 29

0.11

The invention will now be illustrated by the following non-limitingExamples.

EXAMPLES Example 1 Synthesis of Compounds 4-12 (Table I)

To a solution of the substituted benzaldehyde (10 mmol) in 20 mlacetonitrile or ethanol, was added cyanoacetic acid (15 mmol) andpiperidine (10 mmol) and refluxed for 3-8 h at 80° C. Upon thecompletion of the reaction, the above solution was poured into a mixtureof 3M HCl (10 mL) in ice. The solution was stirred for 10 to 15 minutesand the solid was filtered using Buchner funnel. The solid was washedwith water and acetonitrile. The pure compound was obtained uponrecrystallization.

TABLE 2 Elemental Elemental Compound Calculated Observed Yield NumberCompound C % H % N % C % H % N % (%)  4

71.97 8.05 9.33 71.83 8.38 9.35 65  5

73.14 8.59 8.53 72.61 8.37 8.16 70  6

74.12 9.05 7.86 74.18 9.99 7.74 68  7

71.62 6.01 10.44 70.98 6.66 11.36 60  8

72.72 4.58 10.6 70.62 4.7 10.28 62  9

71.97 8.05 9.33 71.48 8.49 9.23 61 10

78.24 5.47 7.60 77.30 5.33 7.88 72 11

69.41 5.82 11.56 68.75 5.50 11.20 68 12

70.29 6.29 10.93 70.51 6.45 11.01 67

Example 2 Synthesis of Compounds 13-21 (Table 3)

To a solution of the substituted o-methoxy-benzaldehyde (10 mmol) in 20ml acetonitrile or ethanol was added cyanoacetic acid (15 mmol) andpiperidine (10 mmol) and refluxed for 10-20 h at 80° C. Upon thecompletion of the reaction, the above solution was poured into a mixtureof 3 M HCl (10 mL) in ice. The solution was stirred for 10 to 15 minutesand the solid was filtered using Buchner funnel. The solid was washedwith water and acetonitrile and the pure compound was obtained uponrecrystallization.

TABLE 3 Elemental Elemental Compound Calculated Observed Yield NumberCompound C % H % N % C % H % N % (%) 13

67.53 7.33 9.26 67.11 7.35 9.14 62 14

69.06 7.93 8.48 70.01 7.66 8.56 60 15

70.36 8.44 7.81 70.51 8.60 7.92 74 16

68.44 6.08 9.39 68.59 6.12 9.48 70 17

69.38 4.79 9.52 69.89 4.99 9.78 66 18

69.06 7.93 8.48 68.95 7.57 8.66 58 19

75.36 5.57 7.03 75.79 5.86 7.34 62 20

74.58 4.90 7.56 74.12 4.47 7.82 66 21

66.16 5.92 10.29 66.36 5.86 10.31 68

Example 3 Synthesis of Compounds 22-29 (Table 4)

To a solution of substituted o-hydroxy-benzaldehyde (10 mmol) in 20 mlethanol, was added diethyl malonate (20 mmol) and piperidine (13 mmol)and refluxed for 8-12 h at 80° C. Upon the completion of the reaction,the above solution was evaporated and further refluxed in 10 ml of 10%NaOH solution. The reaction was quenched with 3M HCl and worked up withethyl acetate. The compound was purified by column chromatography(eluted with 100% ethyl acetate) and recrystallized.

TABLE 4 Elemental Elemental Compound Calculated Observed Yield numberCompound C % H % N % C % H % N % (%) 22

66.42 6.62 4.84 66.54 6.76 4.91 50 23

67.36 5.30 4.91 67.38 5.19 4.98 64 24

68.32 3.94 4.98 68.39 3.97 4.86 53 25

74.79 4.97 3.63 74.87 4.56 3.84 58 26

64.86 5.05 5.40 64.21 4.87 5.12 59 27

68.41 4.07 3.32 68.66 4.34 3.38 55 28

63.45 3.77 3.08 63.55 3.84 2.97 57 29

65.96 4.05 2.96 65.87 3.87 3.08 55

Example 4 Toxicity Study in Healthy ICR Mice

Protocol

60 ICR mice weighing ˜27 grams were randomly assigned into 5 groups (12animals, 6 male and 6 female per group), one group being the controlgroup without any drug administration (food only). The drugs wereadministered twice daily by P.O. or I.P. Group 1 and 3 were administeredorally with 50 mg/Kg of Compound 20 and 18 mg/Kg of Compound 14respectively. Group 2 and 4 mice were injected intraperitoneally with6.67 mg/Kg of Compound 20 and 5.3 mg/Kg of Compound 14 (dissolved inwater) respectively. The body weights of mice were recorded daily for 22days. At the end of the 22^(nd) day, 3 mice were randomly selected fromeach group and their blood samples were collected for detailed analysisof white blood cell (WBC) and red blood cell (RBC) counts. The mice werethen euthanized with CO₂.

Results

TABLE 1 Body Weight Changes of ICR Mice in Compound 20 and Compound 14Toxicity Study Weight in grams Weight in grams Group (Day 0) (Day 22)Compound 20 (50 mg/kg, p.o.) 26.91 32.61 Compound 20 (6.67 mg/kg, i.p)26.98 33.54 Compound 14 (18 mg/kg, p.o) 27.04 32.60 Compound 14 (5.3mg/kg, i.p) 27.05 32.53 No treatment 27.03 35.28

TABLE 2 WBC Count in Toxicity Study of ICR Mice Compound 20 Compound 20Compound 14 Compound 14 No (50 mg/kg, p.o) (6.67 mg/kg, i.p) (18 mg/kg,p.o) (5.3 mg/kg, i.p) treatment WBC ± SEM 4.10 ± 0.3 3.83 ± 0.38 4.57 ±0.49 5.80 ± 0.72 2.97 ± 0.23 (10¹⁰/L)

TABLE 3 REC Count in Toxicity Study of ICR Mice Compound 20 Compound 20Compound 14 Compound 14 No (50 mg/kg, p.o) (6.67 mg/kg, i.p) (18 mg/kg,p.o) (5.3 mg/kg, i.p) treatment REC ± SEM 1.75 ± 0.06 1.82 ± 0.05 1.58 ±0.02 1.58 ± 0.05 1.62 ± 0.09 (10¹³/L)Inference

From this preliminary study, it can be inferred that potent MCT1inhibitors do not cause systemic toxicity and are well tolerated inhealthy ICR mice.

Example 5 Toxicity Study in BALB/C Nude Mice Containing WiDr ColorectalAdenocarcinoma Xenograft

Protocol

Female BALB/c nude mice weighing ˜20 g were taken and 5.0×10⁶ WiDrcancer cells were implanted into their right flanks with a 1:1 mixtureof 0.1 mL PBS and matrigel. The tumor was allowed to grow to a size of˜150 cubic millimeters (14 days). The mice were randomly chosen bychance and assigned into 5 groups (6 mice per group), one group beingthe control group to which tumor was allowed to grow without any drugadministration (food only). The drugs were administered twice daily byP.O. or I.P. Group 1 mice were administered orally with 50 mg/kg ofCompound 20. Groups 2, 3 and 4 mice were treated via IP with 10 mg/kgCompound 20, 8 mg/kg of Compound 14 and 238 mg/kg CHC respectively. Bodyweight changes were recorded every two days for 3 weeks.

Results

TABLE 3 The Body Weight Changes in BALB/C Nude Mice in Toxicity Study ofCompound 20 and Compound 14 Weight in grams Weight in grams Group (Day0) (Day 22) Compound 20 21.85 23.01 (50 mg/kg, p.o.) Compound 20 21.9323.38 (10 mg/kg, i.p.) Compound 14 22.08 23.50 (8 mg/kg, i.p.) Notreatment 22.10 24.21Inference

From this study also, it can be inferred that MCT1 inhibitors do notcause systemic toxicity and are well tolerated in nude mice.

Example 6 Pharmacokinetic Study of Compound 20 in ICR Mice

Protocol:

60 male ICR mice were split into two groups of 30. In each group themice were prescribed Compound 20 at a dosage of 100 mg/kg. This dose wasadministered orally in group 1 and intraperitoneally in group 2. Eachgroup of 30 mice was subdivided into groups of 3 mice. Blood sampleswere taken through orbital sinus method at 0, 0.083, 0.25, 0.5, 1, 2, 4,6, 8 and 24 hours after administering Compound 20 and taken into tubeswith anticoagulant at room temperature at least 30 minutes prior tocentrifugation, then centrifuged at 10000 rpm for 5 minutes at 4° C. toseparate plasma from the blood samples. Following centrifugation, theresulting serum was transferred to clean tubes and stored frozen at −80°C. Immediately following the blood samples, the brain tissue samples ofmice were removed and stored frozen at −80° C.

A 20% brain homogenate was prepared by weighing brain tissue sample andadding, corresponding volume of saline into the manual homogenator. Thedilution factor was 5 (w/v=1 g: 4 mL). The brain tissue sample wasgrinded slowly and repeatedly until it has become homogenous.

Plasma and Brain samples were centrifuged at 15777×g for 5 minutes, and180 ul of supernatant was placed in a 96-well assay plate. To each well,200 ul of a 50% ACN/H₂O was added and the plates were shake-mixed for 3minutes. 5 ul aliquots of each well were then analyzed using LC-MS/MS.

The standard non-compartmental model was used to generate thepharmacokinetic parameters: C_(max), T_(max), t_(1/2), AUC_(0→last),AUC_(0→inf), CL/F, and Vz/F. The maximum plasma concentration, C_(max),and its corresponding time, T_(max) were directly recorded. Eliminationrate constant k_(el) was determined by linear regression of the terminalpoints of the semi-log plot of concentration against time. Eliminationhalf-life was calculated by the formula; t₁₂=0.693/k_(el). The AreaUnder the curve value from time 0 to 24 hours was defined asAUC_(t→last) and calculated using the linear trapezoidal rule.

Results:

Table 4 illustrates the individual and mean plasma concentration data(ng/mL) for Compound 20 after intragastrical administration to mice.

TABLE 4 Plasma Concentration (ng/ml) p.o. (n = 3) Time (h) 1# 2# 3# MeanSD 0 0 0 0 0 0 0.083 2210 4050 4040 3433 1059 0.25 9690 11300 6400 91302498 0.5 19100 20700 6580 15460 7732 1 3610 4830 12700 7047 4934 2 32701380 4790 3147 1708 4 2880 5160 1240 3093 1969 6 1470 2230 3720 24731145 8 5780 2370 3880 4010 1709 24 111 19.3 12.8 47.7 54.9

Table 5 illustrates individual and mean plasma concentration data(ng/mL) for Compound 20 after intraperitoneal administration to mice.

TABLE 5 Plasma Concentration (ng/ml) i.p. (n = 3) Time (h) 1# 2# 3# MeanSD 0 0 0 0 0 0 0.083 18800 33300 40400 30833 11009 0.25 78300 2940060100 55933 24715 0.5 51400 30400 30200 37333 12183 1 31800 45600 2690034767 9697 2 38800 27800 38900 35167 6380 4 25000 24000 26400 25133 12066 14200 3980 5360 7847 5545 8 2760 3160 2590 2837 293 24 41.8 249 70.9121 112

Table 6 illustrates mean pharmacokinetic parameters of Compound 20 inmice,

TABLE 6 PK parameters (Mean ± SD) AUC_(0-last) AUC_(0-∞) t_(1/2Z)T_(max) Vz/F CLz/F C_(max) Dose route (μg/L*h) (μg/L*h) (h) (h) (L/kg)(L/h/kg) (μg/L) p.o. Mean 65740 66011 3.03 0.667 6.55 1.56 17500 SD13050 13365 1.26 0.289 1.81 0.322 4233 i.p. Mean 200796 201474 3.19 0.52.35 0.499 61333 SD 20459 19835 1.16 0.433 1.04 0.0473 16385where AUC_(0-last) is area under the plasma concentration-time curvefrom time zero to time of last measurable concentration, AUC_(0-∞) isarea under the plasma concentration-time curve from time zero toinfinity, T_(max) is time to reach maximum (peak) plasma concentrationfollowing drug administration, T_(1/2) is biological half-life of thedrug, Vz/F is the distribution of a drug between plasma and the rest ofthe body following oral administration, CLz/F is apparent total bodyclearance of the drug from plasma and C_(max) is maximum (peak) plasmadrug concentration.Inference

From this study, it can be inferred that pharmacokinetics parameters forrepresentative Compound 20 are satisfactory.

Example 7 Anticancer Efficacy in Balb/c Nude Mice

Protocol

Female BALB/c nude mice weighing ˜20 g were taken and 5.0×10⁶ WiDrcancer cells were implanted into their right flanks with a 1:1 mixtureof 0.1 mL PBS and matrigel. The tumor was allowed to grow to a size of˜150 cubic millimeters (2 weeks). The mice were randomly chosen bychance and assigned into 5 groups (6 animals/group), one group being thecontrol group to which tumor was allowed to grow without any drugadministration (food only). The drugs were administered twice daily byP.O. or I.P. Group 1 mice were administered orally with 50 mg/kg ofCompound 20. The remaining mice Groups 2, 3 and 4 were treated via IPwith 10 mg/kg Compound 20, 8 mg/kg of Compound 14 and 238 mg/kg CompoundCHC respectively. The tumor volume was recorded every two days using theformulaV=½×a×b ²where ‘a’ is the long diameter of the tumor and ‘b’ is the shortdiameter of the tumor. At the end of the 21^(st) day, the mice wereeuthanized with CO₂ and tumor masses were isolated and weighed. Theinhibition rate was determined using the formula:

${\%\mspace{14mu}{inhibition}} = {\frac{\left( {C - T} \right)}{C} \times 100}$where C is average tumor weight of control group, T is average tumorweight of test group.Results:

The anti-cancer effects of Compound 20, Compound 14, and Compound CHC onthe tumor growth in WiDr xenograft tumor model in Example 6 are shown inFIGS. 1 and 2.

Example 8 Anticancer Efficacy in Balb/c Nude Mice

Protocol:

Female BALB/c nude mice weighing ˜20 g were taken and 5.0×10⁶ WiDrcancer cells were implanted into their right flanks with 1:1 mixture of0.1 mL PBS and matrigel. The mice were arbitrarily chosen by chance andassigned into 7 groups (6 animals/group) and the treatment was startedfrom the same day of the implantation of cancer cells. Mice in group 1were orally administered two times daily with 100 mg/Kg of Compound 20.Mice in group 2 were orally administered four times per day with 50mg/Kg of Compound 20. Group 3 mice were injected intraperitoneally 2times per day with 100 mg/Kg of Compound 20. Group 4 mice were injectedintraperitoneally one time per day with 100 mg/Kg and group 5 mice wereinjected intraperitoneally 2 times per day with 50 mg/Kg of Compound 20.Group 6 was treated with 0.2 mL of 2% DMSO in water intraperitoneally 4times daily and group 7 mice were assigned as the control group to whichtumor was allowed to grow without any drug administration (food only).The tumor volume was recorded every two days using the formulaV=½×a×b ²where a is the long diameter of the tumor and b is the short diameter ofthe tumor. At the end of the 14^(th) day, the mice from group 1, 2, 3, 4and 6 were euthanized with CO₂ and tumor masses were isolated andweighed. The inhibition rate was determined using the formula:

${\%\mspace{14mu}{inhibition}} = {\frac{\left( {C - T} \right)}{C} \times 100}$where C is average tumor weight of control group, T is average tumorweight of test group. The mice from group 5 and 7 were treated with theabove mentioned dosage for one more week (21 days) and were euthanizedwith CO₂ and tumor masses were isolated and weighed. The inhibition ratewas determined using the above formula.Results:

The anti-cancer effect of Compound 20 on the tumor growth in WiDrxenograft tumor model (After 14 Days) from Example 8 are shown in FIG.3.

The anti-cancer effect of Compound 20 on the tumor growth in WiDrxenograft tumor model (After 22 Days) from Example 8 are shown in FIG.4.

The anti-cancer effect of Compound 20 on the tumor growth in WiDrxenograft tumor model from Example 8 are shown in FIG. 5.

The body weight changes in mice from Example 8 are shown in FIG. 6.

Inference

From this data, it can be inferred that Compound 20 can also be used asan adjuvant therapy without much systemic toxicity.

Example 9

The following illustrate representative pharmaceutical dosage forms,containing a compound of formula I (‘Compound X’), for therapeutic orprophylactic use in humans.

(i) Tablet 1 mg/tablet Compound X = 100.0 Lactose 77.5 Povidone 15.0Croscarmellose sodium 12.0 Microcrystalline cellulose 92.5 Magnesiumstearate 3.0 300.0

(ii) Tablet 2 mg/tablet Compound X = 20.0 Microcrystalline cellulose410.0 Starch 50.0 Sodium starch glycolate 15.0 Magnesium stearate 5.0500.0

(iii) Capsule mg/capsule Compound X = 10.0 Colloidal silicon dioxide 1.5Lactose 465.5 Pregelatinized starch 120.0 Magnesium stearate 3.0 600.0

(iv) Injection 1 (1 mg/ml) mg/ml Compound X = (free acid form) 1.0Dibasic sodium phosphate 12.0 Monobasic sodium phosphate 0.7 Sodiumchloride 4.5 1.0N Sodium hydroxide solution q.s. (pH adjustment to7.0-7.5) Water for injection q.s. ad 1 mL

(v) Injection 2 (10 mg/ml) mg/ml Compound X = (free acid form) 10.0Monobasic sodium phosphate 0.3 Dibasic sodium phosphate 1.1 Polyethyleneglycol 400 200.0 1.0N Sodium hydroxide solution q.s. (pH adjustment to7.0-7.5) Water for injection q.s. ad 1 mL

(vi) Aerosol mg/can Compound X = 20.0 Oleic acid 10.0Trichloromonofluoromethane 5,000.0 Dichlorodifluoromethane 10,000.0Dichlorotetrafluoroethane 5,000.0The above formulations may be obtained by conventional procedures wellknown in the pharmaceutical art.

All publications, patents, and patent documents are incorporated byreference herein, as though individually incorporated by reference. Theinvention has been described with reference to various specific andpreferred embodiments and techniques. However, it should be understoodthat many variations and modifications may be made while remainingwithin the spirit and scope of the invention.

What is claimed is:
 1. The compound:

or a salt thereof.
 2. A composition comprising a compound as describedin claim 1, or a pharmaceutically acceptable salt thereof, and apharmaceutically acceptable diluent or carrier.
 3. A method forinhibiting cancer cell growth comprising contacting the cancer cell invitro with an effective amount of a compound as described in claim 1, ora pharmaceutical acceptable salt thereof.